A quantitative model of normal Caenorhabditis elegans embryogenesis and its disruption after stress

Abstract

The invariant lineage of Caenorhabditis elegans has powerful potential for quantifying developmental variability in normal and stressed embryos. Previous studies of division timing by automated lineage tracing suggested that variability in cell cycle timing is low in younger embryos, but manual lineage tracing of specific lineages suggested that variability may increase for later divisions. We developed improved automated lineage tracing methods that allow routine lineage tracing through the last round of embryonic cell divisions and we applied these methods to trace the lineage of 18 wild-type embryos. Cell cycle lengths, division axes and cell positions are remarkably consistent among these embryos at all stages, with only slight increases in variability later in development. The resulting quantitative 4-dimensional model of embryogenesis provides a powerful reference dataset to identify defects in mutants or in embryos that have experienced environmental perturbations. We also traced the lineages of embryos imaged at higher temperatures to quantify the decay in developmental robustness under temperature stress. Developmental variability increases modestly at 25°C compared with 22°C and dramatically at 26°C, and we identify homeotic transformations in a subset of embryos grown at 26°C. The deep lineage tracing methods provide a powerful tool for analysis of normal development, gene expression and mutants and we provide a graphical user interface to allow other researchers to explore the average behavior of arbitrary cells in a reference embryo.

Gene transcription is coordinated with, but not dependent on, cell divisions during C. elegans embryonic fate specification

“…We acquired confocal images with a Leica TCS SP5 (67 z-planes at 0.5 μm spacing and 1.5-minute time spacing) and generated lineages using StarryNite and AceTree as previously described Murray et al, 2006;Murray et al, 2008) but by applying a newer segmentation algorithm (Santella et al, 2010) and higher resolution imaging (Richards et al, 2013). Embryos were mounted in a solution of 20 μm beads in egg buffer/methyl cellulose .…”

Comprehensive single cell-resolution analysis of the role of chromatin regulators in early C. elegans embryogenesis

“…More recently, the Waterston lab pioneered the use of embryos expressing histone::fluorescent protein fusion proteins and nuclear segmentation and tracking algorithms to semi-automatically track nuclear divisions in confocal fluorescent time lapse recordings , greatly improving the number of embryos that can be analyzed compared to manual methods (Hench et al, 2009) and allowing tracking over much longer time windows than can be achieved by image analysis of DIC images (Hamahashi et al, 2005). Others have extended this approach using alternative image analysis methods Giurumescu et al, 2012;Mace et al, 2013;Santella et al, 2010) and, to-date, semi-automated nuclear tracking algorithms have been used to analyze the invariance of cell cycle timing , early (Pohl and Bao, 2010) and late (Giurumescu et al, 2012) cell movements during embryogenesis, gene expression patterns at single cell resolution Murray et al, 2012;Nair et al, 2013), the variability and robustness of development (Moore et al, 2013;Richards et al, 2013), the effects of gene inhibition at single-cell resolution Boeck et al, 2011;Du et al, 2014;Moore et al, 2013), and regulatory interactions and cell fate choices (Du et al, 2014).…”

Infrared laser-induced gene expression for tracking development and function of single C. elegans embryonic neurons

“…Induction of myr-GFP is fastest, followed by myr-mCherry, and Cre-lox-dependent myr-GFP. GFP imaging using a 488 nm laser results in phototoxicity at imaging intensities 5-fold lower than mCherry imaging627. Furthermore, mCherry fluorescence reaches a peak intensity 2.5-fold higher than myr-GFP (Fig.…”

“…As the C. elegans lineage is invariant, descendants of any precursor are known, and their identities can be verified by their terminal morphology, division and migration patterns27 (Fig. 3b,c).…”

“…Nuclear coordinates were taken from a fully lineaged embryo27 at 202 min (AB128) and 357 min (early morphogenesis), and 3D models with coloured sublineages were rendered in MATLAB.…”